The ultimate goal of these studies is to determine the role of intestinal peptide hydrolases in human nutrition and what role, if any, deficiencies of these enzymes play in human diseases. The five peptide hydrolases which have recently been separated from human intestinal brush border will be biochemically characterized and their substrate specificities determined. Protein separating techniques including ion-exchange chromatography, acrylamide gel electrophoresis, starch-gel electrophoresis, and affinity chromatography will be used to isolate each peptide hydrolase present in the cytosol of human intestinal mucosa. Biochemical characteristics and substrate specificity of each isolated enzyme will be determined. Particular emphasis will be placed on determining substrates which are highly discriminating for each brush border and each cytosol enzyme isolated. These substrates will then be used to determine the activity of individual peptide hydrolases in intestinal biopsy specimens from patients with various intestinal diseases including non-tropical sprue, regional enteritis, ulcerative colitis, diarrhea of undetermined etiology, and idiopathic failure to thrive. Results will be compared to controls to determine whether specific peptide hydrolase deficiencies are associated with any of these diseases. One or more of the peptide hydrolases which has been isolated and highly purified from rat or human intestine will be studied in detail in an effort to determine a pattern of substrate specificity which may make it possible to predict the rate of hydrolysis of the hundreds of peptides not readily available for testing. Since defects in intestinal peptide absorption other than peptide hydrolase deficiencies may be a cause of disease, the mechanism(s) of absorption of peptides will be investigated. These studies will be designed to determine the importance of brush border hydrolysis versus intact absorption for various peptides and the role of gamma-glutamyltranspeptidase in transport of free amino acids and intact peptides.